Journal: Journal of Translational Medicine

Article Title: TROP2 confers resistance to oxidative stress-induced cancer cell death through YAP/HMOX1 signaling

doi: 10.1186/s12967-026-07955-z

Figure Lengend Snippet: TROP2 suppression sensitized NSCLC to RSL3-induced ferroptosis in vivo. ( A ) Schematic diagram of the in vivo experimental timeline. Nude mice were subcutaneously injected with control (shCON) or TROP2-knockdown (shTROP2) PC9 cells, followed by intraperitoneal injection of RSL3 (5 mg/kg) or vehicle every other day. ( B ) Representative photographs of dissected subcutaneous tumors from each treatment group at the endpoint of the study. ( C ) Final tumor weights from each group at the endpoint. Data are presented as mean ± SD ( n = 6). ( D ) Tumor growth curves showing tumor volume over time for the four experimental groups. Data are presented as mean ± SEM ( n = 6 mice per group). ( E ) Representative IHC images of tumor sections stained for TROP2, YAP1, HMOX1, ACSL4, and SLC7A11 from the indicated groups. Scale bar, 100 μm. Data are presented as mean ± SD ( n = 6, C) or mean ± SEM ( n = 6 mice per group, D ). Statistical significance for differences among groups was determined by one-way ANOVA followed by Tukey’s post hoc test (C, and for the final time point in D). ∗ P < 0.05, ∗∗ P < 0.01

Article Snippet: After that, the membranes were blocked in 5% fat-free milk for 1 h. The membranes were incubated with the primary antibodies including TROP2 (1:1000, cat #47866, CST), GPX4 (1:1000, cat #AF7020, Beyotime), SLC7A11 (1:1000, cat #AF7992, Beyotime), ACSL4 (1:1000, cat #AG1908, Beyotime), HMOX1 (1:1000, cat #AG2181, Beyotime), YAP1 (1:1000, cat #14074, CST), P-YAP1 (1:1000, cat #13619, CST), LATS1 (1:1000, cat #3477, CST), p-LATS1 (1:1000, cat #9157, CST), Histone3 (1:1000, cat #9715, CST),α-Tubulin (1:1000, cat #2144, CST) at 4 °C overnight and the secondary antibodies at room temperature for 2 h. Protein bands were visualized using the ECL (Bio-Rad, Hercules, CA, USA).

Techniques: In Vivo, Injection, Control, Knockdown, Staining

Journal: Journal of Translational Medicine

Article Title: TROP2 confers resistance to oxidative stress-induced cancer cell death through YAP/HMOX1 signaling

doi: 10.1186/s12967-026-07955-z

Figure Lengend Snippet: TROP2 confers ferroptosis resistance by inactivating YAP to suppress HMOX1 transcription. ( A ) YAP1 mRNA levels in H292 and PC9 cells with TROP2 knockdown, as determined by qRT-PCR. ( B ) YAP1 mRNA levels in PC9 and A549 cells with TROP2 overexpression, as determined by qRT-PCR. ( C ) Representative IF images showing enhanced nuclear localization of YAP (green) in PC9 cells upon TROP2 knockdown. Nuclei were stained with DAPI (blue). Scale bar, 50 μm. ( D ) Representative IF images showing suppressed nuclear localization of YAP (green) in PC9 cells upon TROP2 overexpression. Scale bar, 50 μm. ( E ) Western blot analysis of YAP protein levels in nuclear and cytoplasmic fractions of PC9 and H292 cells with or without TROP2 knockdown. ( F ) Quantitative analysis (bar graph) of the bands was shown adjacent to the representative blots (from E). ( G, H ) Cell viability assessed by CCK-8 assay in PC9 and H292 cells with TROP2 knockdown, with or without concomitant YAP inhibition, following RSL3 treatment. ( I,J ) Measurement of cellular GSH levels in PC9 and H292 cells under the indicated conditions. ( K, L ) Measurement of cellular MDA levels in PC9 and H292 cells under the indicated conditions. Data are presented as mean ± SD ( n = 3). Statistical significance was determined by two-tailed unpaired Student’s t-test ( A , B , F ); two-way ANOVA followed by Sidak’s post hoc test or three-way ANOVA followed by Sidak’s post hoc test ( G - L ). ∗ P < 0.05, ∗∗ P < 0.01

Article Snippet: After that, the membranes were blocked in 5% fat-free milk for 1 h. The membranes were incubated with the primary antibodies including TROP2 (1:1000, cat #47866, CST), GPX4 (1:1000, cat #AF7020, Beyotime), SLC7A11 (1:1000, cat #AF7992, Beyotime), ACSL4 (1:1000, cat #AG1908, Beyotime), HMOX1 (1:1000, cat #AG2181, Beyotime), YAP1 (1:1000, cat #14074, CST), P-YAP1 (1:1000, cat #13619, CST), LATS1 (1:1000, cat #3477, CST), p-LATS1 (1:1000, cat #9157, CST), Histone3 (1:1000, cat #9715, CST),α-Tubulin (1:1000, cat #2144, CST) at 4 °C overnight and the secondary antibodies at room temperature for 2 h. Protein bands were visualized using the ECL (Bio-Rad, Hercules, CA, USA).

Techniques: Knockdown, Quantitative RT-PCR, Over Expression, Staining, Western Blot, CCK-8 Assay, Inhibition, Two Tailed Test

Journal: British journal of cancer

Article Title: Inhibition of the AURKA/YAP1 axis is a promising therapeutic option for overcoming cetuximab resistance in colorectal cancer stem cells.

doi: 10.1038/s41416-024-02649-z

Figure Lengend Snippet: Fig. 1 YAP1 activity correlates with Ser397 phosphorylation during primary resistance to cetuximab in CRC cell lines. a Plot representing the distribution of YAP1 expression in CRC cell lines sensitive to cetuximab compared to the resistant ones ns=non-significant *p < 0.05, two- tailed t-Student’s test. b Plot depicting the distribution of YAP1 activity score in cetuximab-sensitive CRC cell lines compared to the resistant ones. c Correlation analysis between AURKA expression and YAP1 activity score. d Correlation analysis between AURKA and YAP1 expression levels. e Western blot illustrating the basal levels of total amount and phosphorylated YAP1 (Ser397) in CRC cell lines HCA46, SW48 and C10. Tubulin was used as a loading control. Results are plotted as the average ± SD of all the biological replicates and were normalised to the HCA46 cell line (n = 3). ns=non-significant, **p < 0.01, ***p < 0.001, one-way ANOVA. f Gene expression levels of CTGF and CYR61 in CRC cell lines HCA46, SW48 and C10 (n = 3). ns=non-significant, **p < 0.01, ***p < 0.001, one-way ANOVA.

Article Snippet: Primary antibodies used for experiments are the following: ERK (Cell Signaling Technology, #4695, 1:1000), p-ERK (Cell Signaling Technology, #4370, 1:1000), AKT (Cell Signaling Technology, #4691, 1:1000), p-AKT (Santa Cruz Biotechnology, sc-7985-R, 1:250), MET (Proteintech, 25869-1-AP, 1:1000), Tubulin (Proteintech, 66031-1-Ig, 1:3000), YAP1 (Cell Signaling Technology, #14074, 1:1000), p-Ser397-YAP1(Cell Signaling Technology, #13619, 1:1000), MOB-1 (Cell Signaling Technology, #13730, 1:1000), p-MOB-1 (Cell Signaling Technology, #8699 S, 1:1000), LATS-1 (Cell Signaling Technology, #3477,1:1000), p-LATS-1 (Cell Signaling Technology, #8654, 1:1000).

Techniques: Activity Assay, Phospho-proteomics, Expressing, Two Tailed Test, Western Blot, Control, Gene Expression

Journal: British journal of cancer

Article Title: Inhibition of the AURKA/YAP1 axis is a promising therapeutic option for overcoming cetuximab resistance in colorectal cancer stem cells.

doi: 10.1038/s41416-024-02649-z

Figure Lengend Snippet: Fig. 2 AURKA inhibitor alisertib disrupts YAP1 Ser397 phosphorylation and overcomes cetuximab resistance. a Western blots showing the total and phosphorylated forms of YAP1, AKT and ERK after cetuximab and/or alisertib treatment. Cells were stimulated with 40 ng/mL EGF after both treatments to induce ERK and AKT phosphorylation. Tubulin was used as loading control. Results are plotted as the average ± SD of all the biological replicates and were normalised to the EGF condition (n = 3, n = 4 for p-ERK in SW48 cell line).ns=non-significant, **p < 0.01, ***p < 0.001, one-way ANOVA. b Proliferation levels of SW48 and C10 cell lines treated with cetuximab and alisertib, either alone or in combination, relative to control (n = 3). ns=non-significant, **p < 0.01, ***p < 0.001, one-way ANOVA. CTR Control, ALS Alisertib, CTX Cetuximab, COM Combined.

Article Snippet: Primary antibodies used for experiments are the following: ERK (Cell Signaling Technology, #4695, 1:1000), p-ERK (Cell Signaling Technology, #4370, 1:1000), AKT (Cell Signaling Technology, #4691, 1:1000), p-AKT (Santa Cruz Biotechnology, sc-7985-R, 1:250), MET (Proteintech, 25869-1-AP, 1:1000), Tubulin (Proteintech, 66031-1-Ig, 1:3000), YAP1 (Cell Signaling Technology, #14074, 1:1000), p-Ser397-YAP1(Cell Signaling Technology, #13619, 1:1000), MOB-1 (Cell Signaling Technology, #13730, 1:1000), p-MOB-1 (Cell Signaling Technology, #8699 S, 1:1000), LATS-1 (Cell Signaling Technology, #3477,1:1000), p-LATS-1 (Cell Signaling Technology, #8654, 1:1000).

Techniques: Phospho-proteomics, Western Blot, Control

Journal: British journal of cancer

Article Title: Inhibition of the AURKA/YAP1 axis is a promising therapeutic option for overcoming cetuximab resistance in colorectal cancer stem cells.

doi: 10.1038/s41416-024-02649-z

Figure Lengend Snippet: Fig. 5 Alisertib effectively overcomes cetuximab resistance in PDX models with high levles of YAP1 phosphorylation. a Schematic representation outlining the criteria for selecting the tumor for the PDX model based on Western blot screening of YAP1 Ser397 phosphorylation. The tumor with the highest YAP1 phosphorylation levels was chosen for implantation. Mice were treated with 10 mg/kg/day alisertib five days a week and/or 0.4 mg/mice/day cetuximab twice a week for 21 days. Tumor volume and body weight was measured three times a week. Created with BioRender.com b Tumor volume measurement during 21-day treatment with alisertib and/or cetuximab. Representative images of tumors extrated from each group of mice. Results are plotted as the average ± SD of all the tumor volumes for each condition, ***p < 0.001 (n = 6), two-way ANOVA. c T/C (treated/control) ratio of tumors after the 21-day treatment with alisertib, cetuximab and combination. d Ki67 staining of tumors treated with alisertib, cetuximab, the combination of both or neither. *p < 0.05 (n = 3), Welch and Brown-Forsythe ANOVA. CTR Control, ALS Alisertib, CTX Cetuximab, COM Combined.

Article Snippet: Primary antibodies used for experiments are the following: ERK (Cell Signaling Technology, #4695, 1:1000), p-ERK (Cell Signaling Technology, #4370, 1:1000), AKT (Cell Signaling Technology, #4691, 1:1000), p-AKT (Santa Cruz Biotechnology, sc-7985-R, 1:250), MET (Proteintech, 25869-1-AP, 1:1000), Tubulin (Proteintech, 66031-1-Ig, 1:3000), YAP1 (Cell Signaling Technology, #14074, 1:1000), p-Ser397-YAP1(Cell Signaling Technology, #13619, 1:1000), MOB-1 (Cell Signaling Technology, #13730, 1:1000), p-MOB-1 (Cell Signaling Technology, #8699 S, 1:1000), LATS-1 (Cell Signaling Technology, #3477,1:1000), p-LATS-1 (Cell Signaling Technology, #8654, 1:1000).

Techniques: Phospho-proteomics, Western Blot, Control, Staining

Journal: British journal of cancer

Article Title: Inhibition of the AURKA/YAP1 axis is a promising therapeutic option for overcoming cetuximab resistance in colorectal cancer stem cells.

doi: 10.1038/s41416-024-02649-z

Figure Lengend Snippet: Fig. 6 Alisertib reduces c-MET expression levels and impairs CSC features in vivo. a Western blot analysis of total (n = 3) and phosphorylated (n = 4) levels of LATS-1, YAP1, ERK and MOB-1 in tumors treated with alisertib and/or cetuximab, normalized to the control. Results are plotted as the average ± SD of all the biological replicates, **p < 0.01, one-way ANOVA. Differences in p-ERK expression were compared by using Welch and Brown-Forsythe ANOVA since residuals did not meet the homoscedasticity criteria of one-way ANOVA. b Analysis of SOX2 expression levels in tumors after treatment with alisertib and/or cetuximab, normalized to non-treated tumors. Results are plotted as the average ± SD of all the biological replicates, *p < 0.05 (n = 4), one-way ANOVA. c Assessment of ALDH1 activity after treatment with alisertib and/or cetuximab, normalized to non-treated tumors. Results are plotted as the average ± SD of all the biological replicates, *p < 0.05 (n = 4), one-way ANOVA. d. Evaluation of the gene expression levels of SOX2 in tumors after being treated with alisertib and/or cetuximab. Results are plotted as the average ± SD of all the biological replicates, *p < 0.05 (n = 4), one-way ANOVA. CTR Control, ALS Alisertib, CTX Cetuximab, COM Combined.

Article Snippet: Primary antibodies used for experiments are the following: ERK (Cell Signaling Technology, #4695, 1:1000), p-ERK (Cell Signaling Technology, #4370, 1:1000), AKT (Cell Signaling Technology, #4691, 1:1000), p-AKT (Santa Cruz Biotechnology, sc-7985-R, 1:250), MET (Proteintech, 25869-1-AP, 1:1000), Tubulin (Proteintech, 66031-1-Ig, 1:3000), YAP1 (Cell Signaling Technology, #14074, 1:1000), p-Ser397-YAP1(Cell Signaling Technology, #13619, 1:1000), MOB-1 (Cell Signaling Technology, #13730, 1:1000), p-MOB-1 (Cell Signaling Technology, #8699 S, 1:1000), LATS-1 (Cell Signaling Technology, #3477,1:1000), p-LATS-1 (Cell Signaling Technology, #8654, 1:1000).

Techniques: Expressing, In Vivo, Western Blot, Control, Activity Assay, Gene Expression